In higher vertebrates, the glycolytic enzyme enolase (2-phospho-D-glycerate hydrolase; EC 4.2.1.11) is active as a dimer formed from three different subunits, alpha, beta and gamma, encoded by separate genes. The expression of these genes is developmentally regulated in a tissue-specific manner. A shift occurs during development, from the unique embryonic isoform alphaalpha, towards specific isoforms in two tissues with high energy demands: alphagamma and gammagamma in the nervous system, alphabeta and betabeta in striated muscles. The alphaalpha remains widely distributed in adult tissues. Here we report the results of the first extensive study of beta enolase expression during human development. Indeed, the beta subunit is specifically expressed at early stages of human myogenesis. Immunocytochemical analyses demonstrated that it is first detected in the heart of 3-week-old embryos and in the myotomal compartment of somites from 4-week-old embryos. At this stage, the muscle-specific sarcomeric protein titin is expressed in this structure, which will give rise to all body skeletal muscles, but embryonic myosin heavy chain is not yet present. Analyses at the protein level show that, during human ontogenesis, myogenesis is accompanied by an increase in beta enolase expression and by a decrease in the expression of the two other alpha and gamma subunits. Furthermore, beta enolase subunit is expressed in proliferating myoblasts from both embryonic and post-natal muscles. In addition, clonal analysis of primary cell cultures, obtained from the leg muscle of a 7-week-old human embryo, revealed that the beta subunit is present in the dividing myoblasts of all four types, according to the classification of Edom-Vovard et al. [(1999) J Cell Sci 112: 191-199], but not in cells of the non-myogenic lineage. Myoblast fusion is accompanied by a large increase in beta enolase expression. Our results demonstrate that this muscle-specific isoform of a glycolytic enzyme (beta enolase) is among the earliest markers of myogenic differentiation in humans.
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